Most of our projects focus on the function of segments of the two polypeptides that constitute the human hepatic asialoglycoprotein receptor. They are very similar yet exhibit different properties in transfected cells. We will determine the cell-surface distribution of the receptor polypeptides with respect to coated pits, and determine whether ligand or antibody triggers receptor accumulation in coated pits and endocytosis. We showed that when one receptor polypeptide, H2, is expressed without the other, H1, in 3T3 cells, it is synthesized but rapidly degraded. We want to determine whether H2 is degraded in lysosomes, and whether it gets there from the ER without traversing the Golgi. We will make several deletion mutants of H1 and H2, and also chimeras of H1 and H2: by expressing these in 3T3 dells by themselves, or together with normal H1 or H2, we should define segments important in ligand binding, low pH-induced ligand dissociation, endocytosis, transport from ER to Golgi, and rapid degradation. Then we hope to use peptides corresponding to segments of the wild type and mutant proteins to search for cell proteins that bind specifically to that functional segment, and that might be a "transport receptor" or "sorting protein." Other studies focus on regulated transport or newly made hepatic secretory proteins from the ER to the Golgi, emphasizing retinol binding protein and alpha1 antitrypsin. Using 3T3 cells expressing wild type or mutant proteins we want to define the requirements for exit from the ER. We want to identify the putative ER-to-Golgi transport receptor by virtue of its specific binding to the holo, not the apo- form of RBP, and to the wild type and not mutant forms of these proteins. By making and expressing chimeras of RBP and alpha1-antitrypsin, and studying the requirements for their maturation from the ER, we hope to determine whether retention of unfolded proteins in the ER, or active export of folded proteins to the Golgi, is employed.